Identification of plasmon-driven nanoparticle-coalescence-dominated growth of gold nanoplates through nanopore sensing

The fascinating phenomenon that plasmon excitation can convert isotropic silver nanospheres to anisotropic nanoprisms has already been developed into a general synthetic technique since the discovery in 2001. However, the mechanism governing the morphology conversion is described with different reaction processes. So far, the mechanism based on redox reactions dominated anisotropic growth by plasmon-produced hot carriers is widely accepted and developed. Here, we successfully achieved plasmon-driven high yield conversion of gold nanospheres into nanoplates with iodine as the inducer. To investigate the mechanism, nanopore sensing technology is established to statistically study the intermediate species at the single-nanoparticle level. Surprisingly, the morphology conversion is proved as a hot hole-controlled coalescence-dominated growth process. This work conclusively elucidates that a controllable plasmon-driven nanoparticle-coalescence mechanism could enable the production of well-defined anisotropic metal nanostructures and suggests that the nanopore sensing could be of general use for studying the growth process of nanomaterials.

4. On page 16, the authors state that at the beginning of the nanoplate growth reaction "I2 induces the formation of larger Au planar twinned nanoplates through a fast redox reaction." This is then proposed to be followed by coalescence. However, on page 7, the authors also claim that deposition of Au through re-reduction is unlikely because NMR shows no evidence of a change in the citrate concentration. Do the authors have evidence for this first stage of growth where nanoplates form through a redox process? For example, does adding Au ions to the reaction solution change the growth kinetics at this stage?
In addition, how does this two-part mechanism fit with the authors' observations of coalescence of sphere-shaped particles (as opposed to just the coalescence of a plate and a small sphere?) The overall proposed mechanism on page 16 seems somewhat inconsistent with the body of the paper. 5. SiO2 nanoparticles are used as a control to represent the behavior of particles that do not absorb 500 nm light. However, these particles are likely to have different surface functionalization from the Au spheres as well, and therefore may behave differently for that reason also. (The synthesis and/or surface capping of the SiO2 spheres is not described, so it is not clear what is on their surface.) 6. In the abstract, the authors state that "the mechanism based on plasmon-driven hot-electronreduction dominated anisotropic growth…is widely accepted and developed." This is not necessarily true for Ag nanoparticles, as at least one popular mechanism involves the hot-hole-mediated oxidation of citrate, followed by the reduction of Ag+ by the thermalized electrons that result from this oxidation (J. Phys. Chem. C 2007, 111, 8942). 7. In Figure 1, it would be helpful for part (g) to be scaled to the same y-axis as (f) for ease of comparison. The spectra in (g) are somewhat squashed vertically.
Reviewer #2 (Remarks to the Author): Huang et al. reported the translocation events of the intermediate species of nanoparticles during photosynthesis which the morphology conversion was considered as a hot hole-controlled coalescence-dominated growth process. It would be of great importance at single-entity level that the intermediate states could be monitored with high resolutions. However, the methodology used in this work is one of the conventional nanopore detections without further improvements. The insights are not very convincing based on their limited correlation between noisy current recording and limited TEM images, which hinters the impact of the work as well as broader interests. To this end, I would suggest a major revision or transfer to Scientific Report.
1. The reaction condition of photo-induced conversion in Figure 1 is not the same as the rest of nanopore experiments, lacking of 50 mM KNO3 as the electrolyte, which has to be carefully addressed.
2. Although gold nanoparticles do not aggregate at 5 mM or 50 mM KNO3, it will be important to perform dynamic light scattering (DLS) measurements to ensure no aggregation occurred during your storage or upon previous operations.
3. Please provide the diameters, IV curves and baseline RMS noises of all the nanopores used in Figure 2 and 3 as well as in rest of this work.
4. Could you explain the reasons why you used ~20 nm, 1 V for Au nanospheres but ~120 nm, 300 mV for Au nanoplates in Figure 2? Which condition do you use in Figure 3? Could you provide the current at 0h? What are the baseline currents for 0h, 1h, 3h, 5h and 9h in Figure S26? 5. How does the light shine onto your system? Is it focused at the nanopore or passing through the entire trans/cis side of the flowcell? For the latter case, you then measured the diameter evolution more than the light-induced intermediate species, didn't you?
6. It would be critical to understand the min and max hydrodynamic diameters of gold nanoparticles that a nanopore with a certain diameter can cover. Since 9h measurements were performed on 120 nm pores, the cross-section (5 nm/120 nm)^2 should lead to negligible deviations from my point of view. Could you identify the range of particle size your nanopores can detect above your noise level? 7. The reported mean dwell time (Venta et al. Nano Lett. 2014, 14, 9, 5358-5364) of single gold particle translocations is less than 0.1 ms. Could you please make a full description and explains how you make your device work since I am a bit lost in this part? Figure S23. Figure S26 is displayed without detailed experiment conditions. Please make sure the entire piece of work is consistent and well-organized. 9. TEM observations are fantastic. Could you do more statistics based on a decent amount of TEM images to correlate with translocation events?

No scales and units in
Reviewer #3 (Remarks to the Author): The authors present a nice study of their work in supporting a coalescence mechanism as the dominant mechanism of shape transformation of plasmonic nanoparticles. There is a lot of good information within this article to support the proposed mechanism.
1) The authors state that "the mechanism governing the morphology conversion is still elusive, which limits its broader extension". I do not really agree with this. There are two mechanisms at play as proposed in references like #2 and supported by that reference (optical data) and many of the other references used herein. The mechanism is not elusive it has two different components, that are at play in different proportional amounts like with oxygen present, and Ag as the metal the induction period is dominated by Ostwald ripening type processes but reference 2 clearly shows optical signatures for aggregates later in the process when coalescence dominates. The study presented here is not surprisingly dominated by coalescence since the metal is gold which is more easily maintained in a reduced state.
The authors do present a lot of very good data to support this mechanism though.
2) One technical issue I have is with the absorbance spectra. They are repeatedly referred to as UVvis spectra, which is common in literature but absolutely wrong. The y-axis in all of these plots is labelled "Intensity (a.u.)". I think the authors mean Absorbance, which is not a.u. but a unitless quantity and should be written as such. Absorbance is not an intensity measurement, actually it is a lack of intensity.
3) Further to point #2 the absorbance spectra throughout the Supplemental information have absorbances below zero which is not possible these need to be explained at the very least, if there is a background subtraction being used, and probably fixed actually since this data makes no sense.
1. The presence of salt can destabilize citrate-capped gold nanoparticles by screening the surface charge and can lead to aggregation. The authors show in Figure S12 that 5 nm Au spheres are unstable above 10 mM KNO3 while 13 nm Au spheres are unstable above 30 mM KNO3. However, when the authors did their control nanopore experiments to show that the salt conditions of the nanopore experiment don't themselves induce aggregation, they chose to use 13 nm Au spheres for the control instead of 5 nm Au spheres. 13 nm Au spheres are significantly more stable towards aggregation than 5 nm spheres, even in the absence of salt, and therefore this control is not sufficient to conclusively show that the high salt concentrations at the interface of the nanopore do not induce aggregation as an artifact (that would not normally be present during nanoplate growth).

Authors' response:
We are very grateful for the valuable comments. Because welldefined monodispersed 13 nm Au nanoparticles were easily obtained, they were used for control experiment to demonstrate the aggregation is not artifact caused by the detection method. The control experiment with 13 nm is also very meaningful. The TEM (Supplementary Figure 9) results show that the size of Au nanoparticles is already close to or more than 13 nm after 30 min of growth reaction (Fig. R1). Therefore, nanopore analysis using 13 nm Au nanoparticles as the control experiment can exclude the artifact during conversion process (over 30 min).
To make a more convincing control experiment for the smaller size Au nanoparticles, we tried to obtain monodisperse Au seed nanoparticles by centrifugation to get rid of most of 2 large aggregations. As shown in Fig. R2, Au nanoparticles with small size and much better monodispersity were obtained. The relative translocation events are mainly single peak events (more than 90%), indicating that the salt concentration gradient does not induce aggregation as an artifact.   Figure S 20 rearrange to form a plate shape? It is well-known that 5 nm Au spheres aggregate and coalesce into larger particles (this is why the 5 nm seeds 3 used for seed-mediated syntheses must be made fresh daily), and therefore it would be helpful to see more evidence of how this aggregation directly correlates with nanoplate formation.

Authors' response:
We are very grateful for the valuable comments. Taking advantage of TEM tracking and nanopore analysis, it is demonstrated that aggregations happen during the transformation of Au nanospheres into nanoplates in our system.
To prove that the aggregation of Au nanoparticles can make nanoplate, it is necessary to show that the aggregation and coalescence process can produce planar twinned Au nanostructures, which can develop to Au nanoplates. Here, we utilize the TEM tracking strategy to get new evidences. As shown in the Fig. R3, we traced several linear aggregates using TEM before and after 30 min reaction in the real condition, and found that the aggregate coalesced into one nanostructure. For Fig. R3c, the insert HRTEM image clearly show that the linear aggregate changed to a planar twinned nanostructure, because the twin plan (highlighted by the white dash line) is just vertical to the TEM chip. It is not always easy to identify all of these twinned structures with different postures by TEM. Thus, we further grow these nanoparticles with previous reported method (details shown in the figure legend), by which the planar twinned Au seeds will develop to Au nanoplates and other twinned Au seeds (mainly penta twinned structure) will grow to nanospheres (Nat. 3. The authors show examples of a single particle merging into a larger plate structure as well as long chains of aggregated spheres. Some analysis of the reaction order might provide evidence into whether the process of plate formation involves stepwise addition of smaller spheres to larger particles or the concerted merging of many particles at once. The reaction order for plasmon-mediated Ag nanoprism formation has previously been presented as one piece of evidence against the oriented attachment hypothesis for that particular system (J. Am. Chem. Soc., 2008, 130, 9500).

Authors' response:
We are very grateful for the valuable comments. According to our observations, the linear aggregation of Au nanospheres can merge together fast to form planar twinned nanostructure in the reaction condition, as shown in the Fig. R3. In the initial stage, we have not observed the intermediate structures with one nanoplate merging with 5 many nanoparticles at one time. Thus, the stepwise aggregation may exist as the reviewer mentioned. It is proposed that the Au nanoplate mainly form with coalescence of Au nanoparticles one by one. However, when the Au nanoplate grow in size. There are opportunities for the coalescence of more Au nanoparticles on one Au nanoplate (Fig. R4).
Based on the evidences from TEM tracking and nanopore analysis, the plasmondriven conversion of Au nanospheres to nanoplates indeed follows a different mechanism towards most reports for the silver counterpart, which is proposed mainly following the oxidation and re-reduction process. 4. On page 16, the authors state that at the beginning of the nanoplate growth reaction "I2 induces the formation of larger Au planar twinned nanoplates through a fast redox reaction." This is then proposed to be followed by coalescence. However, on page 7, the authors also claim that deposition of Au through re-reduction is unlikely because NMR shows no evidence of a change in the citrate concentration. Do the authors have evidence for this first stage of growth where nanoplates form through a redox process? For example, does adding Au ions to the reaction solution change the growth kinetics at this stage? In addition, how does this two-part mechanism fit with the authors' observations of coalescence of sphere-shaped particles (as opposed to just the coalescence of a plate and a small sphere?) The overall proposed mechanism on page 16 seems somewhat inconsistent with the body of the paper.

Authors' response:
We are very grateful for the valuable comments. In the previous manuscript, the growth process is described as two parts: fast redox reaction and slow coalescence-dominated growth. For the coalescence-dominated growth, strong evidences can demonstrate this process. For example, NMR shows no obvious change was found in the concentration of citrate; intermediate aggregates were characterized by TEM tracking and nanopore detection. Since we did not have solid evidence that the nanoparticlecoalescence can produce planar twinned nanostructures then, we proposed the redoxdominated process for the very beginning stage, which is quite similar to the reported mechanism of silver system. However, in the revised manuscript, according to new evidences, we have got a more comprehensive description for the growth process. It is demonstrated that the aggregation and coalescence can produce planar twinned Au nanostructures. Thus, the nanoparticlecoalescence growth pathway is proposed to dominate for the whole growth process.
At the beginning of the reaction, the adding of I2 can selectively etch the Au 6 nanoparticles (Supplementary Figure 6). It is still possible that the released Au ions can be re-reduced and deposited onto more stable Au nanoparticles. The citrate can act as reducing agent, but the consumption amount is too small to be clearly detected by NMR.
To prove it, as the reviewer suggested, we added additional HAuCl4 in the solution. As shown in Fig. R5, the stronger extinction peaks exhibit the growth of the Au nanostructures. However, no obvious dominated SPR peak of larger Au nanoplates appeared for the sample with additional 100 ul HAuCl4 (10 mM) after 180 min light irradiation. It indicates that the addition of Au ions in the conversion solution is not helpful for the selective generation of Au nanoplates. We have updated the growth mechanism in the revised manuscript, in which the nanoparticle-coalescence growth dominates in the whole growth process. 5. SiO2 nanoparticles are used as a control to represent the behavior of particles that do not absorb 500 nm light. However, these particles are likely to have different surface functionalization from the Au spheres as well, and therefore may behave differently for that reason also. (The synthesis and/or surface capping of the SiO2 spheres is not described, so it is not clear what is on their surface.)

Authors' response:
We are very grateful for the valuable comments. The SiO2 nanoparticles were prepared as reported method (J. Am. Chem. Soc., 2012, 134(13): 5722-5725.), which is added in the revised manuscript. The surface of SiO2 was mainly 7 capped by CTAC. Because the SiO2 nanoparticles do not absorb 500 nm light, the purpose of the control experiment with SiO2 nanoparticles is to demonstrate that the change of translocation events through nanopore is caused by the absorption of light by the Au nanoparticles, not other effects that causing changes to the nanopore or solution. Because the surfactants (such as citrate and CTAC) do not absorb 500 nm light, the plasmon excitation of Au nanoparticle should play the main role as we discussed in the manuscript.
6. In the abstract, the authors state that "the mechanism based on plasmon-driven hotelectron-reduction dominated anisotropic growth…is widely accepted and developed." This is not necessarily true for Ag nanoparticles, as at least one popular mechanism involves the hot-hole-mediated oxidation of citrate, followed by the reduction of Ag + by the thermalized electrons that result from this oxidation (J. Phys. Chem. C., 2007, 111, 8942).

Authors' response:
We are very grateful for the valuable comments. We have changed these sentences accordingly.
7. In Figure 1, it would be helpful for part (g) to be scaled to the same y-axis as (f) for ease of comparison. The spectra in (g) are somewhat squashed vertically.

Authors' response:
We are very grateful for the valuable comments. We have plotted Fig.  1g as suggested by the reviewer. 8

Reviewer #2 (Remarks to the Author):
Huang et al. reported the translocation events of the intermediate species of nanoparticles during photosynthesis which the morphology conversion was considered as a hot holecontrolled coalescence-dominated growth process. It would be of great importance at single-entity level that the intermediate states could be monitored with high resolutions. However, the methodology used in this work is one of the conventional nanopore detections without further improvements. The insights are not very convincing based on their limited correlation between noisy current recording and limited TEM images, which hinters the impact of the work as well as broader interests. To this end, I would suggest a major revision or transfer to Scientific Report.

Authors' response:
We are very grateful for the valuable comments. In this work, we observed the direct plasmon-driven morphology conversion from Au nanospheres to Au nanoplates, which is a very interesting process. To confirm the growth way, we mainly utilized both TEM tracking and nanopore detection to study the intermediate species. The main contribution of nanopore sensing technology is that it plays a very important role in confirming intermediate aggregated states at single-entity level in the solution, while there are significant limitations to confirm it under TEM characterization (even with liquid-cell TEM).
Additionally, as a powerful tool, we also demonstrated that the nanopore sensing could be of general use for studying the growth mechanism of nanostructures. The salt concentration gradient strategy has been used for the research of DNA and proteins, we demonstrated that it is indeed helpful for nanopore analysis of metal nanoparticles, improving nanoparticle capture rates and translation time resolution. We present here the strategy of determining the shape and coalescence of nanoparticle in solution using nanopore sensing.
In the revised manuscript, we strengthened the studies of the capability of nanopore detection and TEM tracking for the nanoparticle conversion. We believe that this revised manuscript will be of great interest to researchers working on better understanding plasmon-mediated syntheses and studying the nanostructures shape evolution. Figure 1 is not the same as the rest of nanopore experiments, lacking of 50 mM KNO3 as the electrolyte, which has to be carefully addressed.

Authors' response:
We are very grateful for the valuable comments. For the nanopore characterization, certain amount of growth solution is taken out and KNO3 electrolyte was added to support the electrochemical analysis. It is demonstrated that the present of 5mM KNO3 will not cause aggregation of Au nanoparticles, which is good for capturing the intermediate species by nanopore sensing. Additionally, no light irradiation was supplied during the morphology characterizations with nanopore. With nanopore sensing, we did not directly monitor the evolution of nanostructures, but captured the intermediates for different reaction time. 9 2. Although gold nanoparticles do not aggregate at 5 mM or 50 mM KNO3, it will be important to perform dynamic light scattering (DLS) measurements to ensure no aggregation occurred during your storage or upon previous operations.
Authors' response: Thank you for your valuable suggestions. We have supplemented time-dependent DLS of initial Au seeds (Fig. R6). The dominant size distribution shows no significant aggregation of seeds in 5 mM KNO3 solution during one hour. It suggests that 5 mM KNO3 does not lead to particle aggregation during nanopore testing. Combined with the control experiments of nanopore characterizations with monodispersed ~5 nm and ~13 nm Au nanoparticles (Supplementary Figure 21 and Figure 22), we further confirm the advantages of the nanopore sensing strategy without artifact.  4. Could you explain the reasons why you used ~20 nm, 1 V for Au nanospheres but ~120 nm, 300 mV for Au nanoplates in Figure 2?
Authors' response: We are very grateful for the valuable comments. The nanopore size was chosen by the size distribution of nanostructures from TEM analysis to ensure that it can cover the range of the particle size. The voltage was chosen to ensure a high event rate and to allow the nanopore to work for a long time without blocking. For example, as shown in the Fig. R8, when the applied voltage is 400 mV during the tests of Au nanoplates, obvious blockage occurs after ~200 s. Therefore, it is very important to select appropriate conditions to obtain nanopore signals with high signal-to-noise ratios and high event rates.  5. How does the light shine onto your system? Is it focused at the nanopore or passing through the entire trans/cis side of the flowcell? For the latter case, you then measured the diameter evolution more than the light-induced intermediate species, didn't you?
Authors' response: We are very grateful for the valuable comments. During nanopore analysis of the shape of the Au nanoparticles, no light shined on the system. Thus, intermediate species were captured for detection. Only for studying the effect of light irradiation on Au nanoparticles, the light was used and passed through the entire solution.
6. It would be critical to understand the min and max hydrodynamic diameters of gold nanoparticles that a nanopore with a certain diameter can cover. Since 9h measurements were performed on 120 nm pores, the cross-section (5 nm/120 nm)^2 should lead to negligible deviations from my point of view. Could you identify the range of particle size your nanopores can detect above your noise level?

Authors' response:
We are very grateful for the valuable comments. We totally agree with the reviewer that it is important whether the nanopore we choose can cover the nanoparticles in the sample. The sample with the longest reaction time used for nanopore characterization is from 9h reaction. As shown in Fig. R10, the size distribution of nanoplates in the 9 h sample is around 100 nm, while the nanoparticles in the sample are widely distributed with a minimum of about 35 nm (not 5 nm seeds). We further verified the translocation of 35 nm nanoparticles using a 120 nm nanopore with a voltage of 300 mV. Distinct nanoparticle translocation events can be detected. Therefore, we believe that it could match the nanoparticle distribution range of the 9 h sample using a 120 nm nanopore.  Lett. 2014, 14, 9, 5358-5364) of single gold particle translocations is less than 0.1 ms. Could you please make a full description and explains how you make your device work since I am a bit lost in this part?
Authors' response: We are very grateful for the valuable comments. We suppose that this difference may be caused by the method of salt gradient and the conical nanopipette nanopore used in this work, while a 40 nm-thick SiNx nanopore was used in the 1 mM to 100 mM KCl solution for both cis and trans sides in the mentioned report (Nano Lett. 2014, 14, 9, 5358-5364). As we described in Supplementary Figure 14 the conical nanopore could introduce a reverse flow by a salt concentration gradient (induced reverse 13 electroosmotic flow, IREOF). When the Au nanoparticles or nanoplates are added into the low salt concentration, it is easier to be captured by the inlet flow (e.g., IREOF) near the nanopore (Anal. Chem., 2016, 88, 9251−9258). Then, the Au nanoparticle or nanoplate translocated to the high concentration, it would be retarded by the stronger electroosmosis force (the direction of the electroosmosis force is opposite to the sample movement), resulting in taking the longer time pass through the nanopore (Biophys. J., 2013, 105,  776−782; Nat. Nanotechnol., 2010, 5, 160−165). Therefore, the dwell time of Au nanoparticles translations is longer in this work. Fig. R11. Schematic illustration of a conical nanopore with a salt gradient for nanoparticles translocation. Figure S 23. Figure S 26 is displayed without detailed experiment conditions. Please make sure the entire piece of work is consistent and well-organized.

Authors' response:
We are very grateful for the valuable comments. We have revised these figures, and double-checked and revised the manuscript to avoid such problems. 9. TEM observations are fantastic. Could you do more statistics based on a decent amount of TEM images to correlate with translocation events?
Authors' response: We are very grateful for the valuable comments. It would be more informative to combine the statistics of all TEM images with the results of the translocation events. In this work, we mainly utilized the nanopore sensing technique to confirm the aggregation of nanoparticles and calculate the percentage of the Au nanoplates and Au nanospheres for different reaction time. As shown in Fig. R10, for the 9 h sample, the counting of the percentage of Au nanoplates (~88.09%) and Au nanospheres (~11.91%) are close to the statistical analysis from nanopore sensing results (~ 85.98% for nanoplates; ~ 14.02% for nanospheres). For the 5 h sample (as shown in Fig. R12), the counting of the percentage of Au nanoplates (~60.38%) and Au nanospheres (~39.62%) are also close to the statistical analysis from nanopore sensing results (~ 51.83% for nanoplates; ~ 48.17% for nanospheres). However, we encountered problems to do the similar calculations for samples from shorter time reactions, such as 3 h sample (as shown in Fig. R13). It is hard to identify the Au nanoplates from the Au nanospheres when their sizes are close, such as the Au nanostructures marked with red circles in Fig. R13a and 14a. The nanopore analysis exhibits much better capability to identify the Au nanoplate from the special shape of the translocation events (~41.45% for nanoplates; ~58.55% for nanospheres), and show higher percentage of Au nanoplates compared with the counting result from TEM images (~23.94% for nanoplates; ~76.06% for nanospheres). Obviously, certain amount of Au nanoplates is treated as Au nanospheres in the TEM images. Another challenge is that it is hard to identify the aggregates (e.g.,1 h sample, Fig. R14) from the aggregation artifacts caused by the TEM sample preparation. The subjective judgement for the nanostructures from TEM may cause large deviation for the results. As shown in Fig. R14b, there is a large difference between TEM results and nanopore results in the 1 h sample statistics. Nanoplates only accounted for 9.62% from TEM results. However, the content of nanoplates in nanopore events reached 31.81%. However, for the well-dispersed 13 nm nanoparticles (Fig. R15a) and pure nanoplates (Fig. R10c), the results of TEM statistics and the nanopore translocation events can be correlated well with each other.
Overall, in this system, the statistics analysis from TEM is not suitable to supply correct information for the aggregation states and the calculation of the percentage of Au nanostructures in the early stage. Instead, the nanopore sensing and analysis is very helpful to supply these statistics information. However, the TEM tracking method plays very important role for studying the growth process, as we discussed in the manuscript and responses.

Reviewer #3 (Remarks to the Author):
The authors present a nice study of their work in supporting a coalescence mechanism as the dominant mechanism of shape transformation of plasmonic nanoparticles. There is a lot of good information within this article to support the proposed mechanism. 1) The authors state that "the mechanism governing the morphology conversion is still elusive, which limits its broader extension". I do not really agree with this. There are two mechanisms at play as proposed in references like #2 and supported by that reference (optical data) and many of the other references used herein. The mechanism is not elusive it has two different components, that are at play in different proportional amounts like with oxygen present, and Ag as the metal the induction period is dominated by Ostwald ripening type processes but reference 2 clearly shows optical signatures for aggregates later in the process when coalescence dominates.

Authors' response:
We are very grateful for the valuable comments. We agree that the sentence "the mechanism governing the morphology conversion is still elusive, which limits its broader extension" was not an accurate description. We have changed it in the revised manuscript.
The study presented here is not surprisingly dominated by coalescence since the metal is gold which is more easily maintained in a reduced state. The authors do present a lot of very good data to support this mechanism though.
2) One technical issue I have is with the absorbance spectra. They are repeatedly referred to as UV-vis spectra, which is common in literature but absolutely wrong. The y-axis in all of these plots is labelled "Intensity (a.u.)". I think the authors mean Absorbance, which is not a.u. but a unitless quantity and should be written as such. Absorbance is not an intensity measurement, actually it is a lack of intensity.

Authors' response:
We are very grateful for the valuable comments. In this article, nanoparticle solutions are usually characterized by measuring their extinction spectra with a UV-vis spectrometer. Although sometimes referred to as absorption spectra, extinction is actually measured as the sum of absorption and scattering of light by the nanoparticles. Absorption dominates for small nanoparticles, and scattering for large ones, but for intermediate sizes (in the range of 40−100 nm), absorption and scattering can be of a similar order of magnitude (Craig F. Bohren and Donald R. Huffman. Absorption and scattering of light by small particles. John Wiley & Sons, 2008.). Therefore, we have now updated UV-vis graphs in the revised manuscript using extinction for all nanoparticles and absorbance for solution without a unit quantity.
3) Further to point #2 the absorbance spectra throughout the Supplemental information have absorbances below zero which is not possible these need to be explained at the very least, if there is a background subtraction being used, and probably fixed actually since this data makes no sense.

Authors' response:
We are very grateful for the valuable comments. There should be a wrong background subtraction for the absorbance below zero, supplemental Fig. 5a, 7b, and 8a, which were supplemented. We have collected new data and updated them in the revised manuscript. They do not change the findings or interpretation in the text. Thanks very much for pointing out this mistake.
The authors have addressed my concerns.
Reviewer #2 (Remarks to the Author): The authors did a lot of efforts on the improvements of the quality of this work which I sincerely appreciate. However, I become even less convinced at this stage by their fragmentations of nanopore translocation evidences by applying a 5 kHz filter (I guess a low-pass filter) to identify the events. The translocation experiments were performed by transferring some of the reacted solutions out of the bulk into the nanopore device, which is definitely misled in the abstract of the striking phrase "during reaction". Here are the details which I suggest the authors to re-organize them indepth. Otherwise, a rejection or transferring to other journals may not be a bad idea.
1. For Q4 R2, leaving all the scatter plots in the Supporting information is definitely not great. I like Figure S27, S28 and S31, however, some of them should be in the main text but not some of the representative events instead. Moreover, these figures are not at the same scale which disappoint me once more.
(1) When you use different sizes of nanopores for the reacted solutions stopped at different time, smaller aggregations will be ignored by larger pore sizes. How could you compare the degree of your aggregations?
(2) Since you have applied a 5 kHz filter (I guess a low-pass filter) to identify the events, then the events faster than 0.2 ms in a lot of your measurements (i.e. Figure S16, S21b, S22d to name a few) cannot be trustable at all. Although the event ratio seems correlated with TEM observation but once you have all the events below 0.2 ms removed, what is going to happen?
(3) Was Figure S31a without light illumination the same as Figure S16? Then what is the pie chart with 500nm illumination going to be?
(4) For 5 nm gold spheres the average translocation time of 0.372 ms was longer than that (0.1 ms) of 13 nm gold spheres in Figure S21b. Is this reasonable? Updated Fig. 3. Nanopore analysis of the morphological conversion mechanism of Au nanospheres to Au nanoplates. a, The ratio of nanoplates and coalescence in the process of morphology transformation obtained from CNN prediction model and peak analysis code, respectively. 1135, 1035, 1283 and 1198 events were obtained in the 1 h, 3 h, 5 h and 9 h samples, respectively. A representative current trace and event scatter plots of dwell time vs. current blockade of all the experiments are displayed in Supplementary  Fig. 27 and 28. b, Typical nanopore current-time peak shape for nanoparticle-nanoparticle, nanoparticle-nanoplate and nanoplate-nanoplate coalescence events (left to right). c, Current versus time trace of Au nanospheres without (left) and with 500 nm light (right). d, Scatter plots for td vs. Ib of Au nanospheres translocation without (left) and with 500 nm light (right) from c, and along the sides are the corresponding histograms. Measurements were collected using a same ~20 nm nanopore at a bias voltage of 1 V. The baseline current is about 310 pA and RMS is 2.18 pA. e, Box plots of event rate and Ib/I0 from Au nanospheres with 500 nm light and without light. Box plots show median (black line), mean (black square box), quartiles (boxes) and range (whiskers). Mann-Whitney U tests, *** p < 0.001.
For the analysis of the events, we have tuned the scale the same (updated Fig. S28 and S31) as suggested by the reviewer. Only for the data from 9h, a larger scale has to be used because the values are much larger.
Additionally, we would like to emphasize that the analysis based on scatter plots of analyte (td vs. Ib) are no longer working well in distinguishing Au nanoparticles, nanoplates and their aggregates in this system. Actually, we focused on the peak shape of the events rather than the translocation time and blocking current.
(1) When you use different sizes of nanopores for the reacted solutions stopped at different time, smaller aggregations will be ignored by larger pore sizes. How could you compare the degree of your aggregations? Authors' response: We are very grateful for the comments. As we mentioned before, nanoparticles are always aggregating throughout the formation of nanoplates. Thus, the smallest aggregates should be at the beginning of the reaction, and the ~ 20 nm nanopores we used can completely cover the diameter of the aggregates at this stage. After 9 h of reaction, the size difference between nanoparticles and nanoplates in the aggregates is the largest. As we have demonstrated, the diameter of the smallest nanoparticle in the sample is about 35 nm, and we can detect these nanoparticle translocation events using a 120 nm nanopore. The nanopore size was chosen by the size distribution of nanostructures from TEM analysis to ensure that it can cover the range of the particle size.  It is worth to mention that there may be deviation for the percentage analysis caused by certain aggregations with less anisotropic morphologies. For example, as shown in the Fig. R2, these aggregates may not produce a distinct two-peak shape during nanopore detection, because the size difference of the two domains are too large or the coalescence is almost finished. In this case, the degree of detected aggregations may be lower than the real status. To reduce the deviation, we always collected more than 1000 events. Nevertheless, the deviation will not change the conclusion of the coalescence-induced growth of Au nanoplates.
(2) Since you have applied a 5 kHz filter (I guess a low-pass filter) to identify the events, then the events faster than 0.2 ms in a lot of your measurements (i.e. Figure S16, S21b, S22d to name a few) cannot be trustable at all. Although the event ratio seems correlated with TEM observation but once you have all the events below 0.2 ms removed, what is going to happen?